Early embryo biopsy, "stem cells" - and Twins

Dianne N. Irving
Copyright November 11, 2005
Reproduced with Permission

I. Introduction

Great excitement was recently generated about new research studies involving biopsied early human embryos as a new source of human embryonic "stem cells", a source that would by-pass the ethical objections of other "alternative methods" being discussed lately:

The most basic objection to human embryonic stem (ES) cell research is rooted in the fact that ES cell derivation deprives embryos of any further potential to develop into a complete human being. ES cell lines are conventionally isolated from the inner cell mass of blastocysts and, in a few instances, from cleavage stage embryos. So far, there have been no reports in the literature of stem cell lines derived using an approach that does not require embryo destruction. Here we report an alternative method of establishing ES cell lines - using a technique of single-cell embryo biopsy similar to that used in pre-implantation genetic diagnosis of genetic defects - that does not interfere with the developmental potential of embryos. .. The ability to generate human ES cells without the destruction of ex utero embryos would reduce or eliminate the ethical concerns of many.1 (emphases added)

But these scientific findings are not essentially new or unexpected; nor do they "reduce or eliminate the ethical concerns of many". Or perhaps the authors are still unfamiliar with both natural and artificial monozygotic twinning?

What is finally talked about in this article is the well-known scientific fact that many early human embryos diagnosed as "damaged" (e.g., after freezing and thawing), or that undergo prenatal genetic diagnosis, could heal themselves, be implanted, and be born normal.2 What is also talked about is that the separated cells (blastomeres) biopsied or separated from the whole early human embryo can go on to produce human embryonic "stem cell" lines.

But what is still not talked about is the fact that these cells (blastomeres) isolated by biopsy can also revert to new embryos in vitro - and then be used as a continuous source of new human "biological" materials for research, or be implanted, allowed to develop to term, and be born. This is essentially what is already being done in IVF clinics when clinicians artificially "twin" (clone) an early human embryo for "infertility treatments". Even in natural monozygotic twinning in utero, the separated isolated cell/s (blastomere/s) revert to new embryos, develop, and are born.

II. Twinning

But no one wants to talk about "twinning". Why? Because then they will also have to explain the science behind monozygotic twinning, especially the natural process of "regulation" and the totipotency (not pluripotency) of these early human embryonic cells (blastomeres). If the public or even other researchers understood these scientific facts they might begin to have second thoughts about a lot of things. This calls into question whether or not these new alternative methods are for real, or simply new clever ways to camophlage "scientifically" what is really going on in the labs.

It is precisely the natural biological process of "regulation" and the "totipotent" properties of all early human embryonic "stem cells" which explain to a great extent what makes possible such early embryonic "healing" as reported in this recent study, as well as explain the reversion of early embryonic cells to new embryos.

It has been well known for decades that "regulation" is in large part responsible for both the healing of damaged early human embryos as well as for the reversion of their separated totipotent cells (blastomeres) to new human embryos.3 Indeed, "regulation" has been routinely used to explain the natural formation of monozygotic twins in utero in human embryology textbooks for decades,4 and explains as well the use in IVF clinics of this kind of cloning technique to artificially produce born monozygotic "twins" and "triplets" (called "blastomere separation" and "blastocyst splitting").5

And another thing not talked about is that because these human embryonic cells (blastomeres) that are "biopsied" from early human embryos are totipotent (not "pluripotent"),6 they can also revert to new human embryos in vitro and develop in vitro to the blastocyst stage:

To investigate the development potential of blastomeres isolated from 4-cell human embryos. METHODS: Human oocytes were collected after controlled ovarian hyperstimulation and fertilized in vitro. The zona pellucida of 4-cell stage embryos were removed and the embryos were separated to single blastomeres and cultured in vitro. The development and differentiation of the blastomeres were observed. RESULTS: 83 blastomeres were isolated from 20 4-cell and 2 3-cell human embryos. Of those, 35 developed to expanded blastocyst. CONCLUSION: Blastomeres isolated from the 4-cell human embryos have the competence to develop to expanded blastocyst.7 (emphases added)

The same has been documented in studies with other animals.8

Indeed, isolated cells (blastomeres) from early embryos can undergo regulation in vitro to form blastocysts, which when implanted develop to term and result in normal births (an artificial way to produce what happens in natural in utero monozygotic twinning):

The aim of this study was to obtain mice, hopefully identical multiplets, from single diploid blastomeres isolated at the 4-cell stage, or from pairs of sister blastomeres isolated at the 8-cell stage. ... The aggregates were cultured in vitro up to the blastocyst stage. Each quartet (occasionally triplet or doublet) of chimaeric blastocysts was transplanted to the oviduct of a separate pseudopregnant recipient. ... These observations strongly suggest that ... the described method can be used in future to produce twins, triplets and quadruplets in the mouse. Our study confirms earlier work by Kelly (1975, 1977) that 'quarter' blastomeres of the mouse are still totipotent.9 (emphases added)

III. Ethical Concerns Remain

Not only are IVF and similar ART procedures per se still ethically debatable, the scientific fact that isolated human blastomeres from biopsied early human embryos have the natural capacity to revert to new human embryos in vitro is also ethically significant. Such cells could obviously provide a constant supply of both normal and abnormal embryos (human beings) for research purposes only - including research involving all kinds of human cloning, human nanotechnology, human genetic engineering, etc. All such research necessarily involves their mutilation and destruction.

Such experimental totipotent human cells and experimental human embryos could also be implanted, further researched upon in utero, and allowed or not allowed to develop to birth. Women undergoing such experimental "infertility" procedures or "donating" their human embryos would not be able to give ethically or legally valid informed consent without understanding "what" it is that they are donating or is being implanted into their wombs - nor understand all of the possible reasons why they have so many spontaneous abortions.

Professional medical and research ethics and standards - if they still or ever existed - should require the use and reporting of all accurate scientific facts involved in these studies by the researchers, including essential biological processes involved such as "regulation" and "totipotency" which scientifically explain and support these findings. Respected scientific/medical journals should likewise require such disclosure and scientific explantations before agreeing to publish these studies, as well as continue to require "conflict of interest" statements of the authors.

Finally, politicians, legislators and the public have the right to be fully - not partially, conveniently, or deceptively - informed about the accurate scientific facts involved in all these related issues that they are increasingly required to make decisions about. Without such accurate facts there is no truly democratic process at all.

Note - note one word about "religion", "theology", "belief systems", "opinions", or "prolife" is needed for the above scientific or ethical analysis. Just the facts.


1 Young Chung, Irina Klimanskaya, Sandy Becker, Joel Marh, Shi-Jiang Lu, Julie Johnson, Lorraine Meisner and Robert Lanza, "Embryonic and extraembryonic stem cell lines derived from single mouse blastomeres", Nature 16 October 2005 | doi: 10.1038/nature04277, at: http://www.nature.com/nature/journal/vaop/ncurrent/abs/nature04277.html. As noted in the full article: The authors declare competing financial interests. Declaration: Y.C., I.K., S.B., J.M., S.-J.L. and R.L. are employees of Advanced Cell Technology, a biotechnology company in the field of regenerative medicine. J.J. and L.M. declare no conflict of interest. [Back]

2 E.g., see Ronan O'Rahilly and Fabiola Muller, Human Embryology & Teratology (New York: Wiley-Liss, 2001): "Biopsy of an embryo can be performed by removing one cell from a 4-cell, or two cells from an 8-cell, embryo. This does not seem to decrease the developmental capacity of the remaining cells." (p. 37) Kay T. Elder, "Laboratory techniques: Oocyte collection and embryo culture" in Peter Brinsden (ed.), A Textbook of in vitro Fertilization and Assisted Reproduction, 2nd edition (New York: The Parthenon Publishing Group, 1999): "Surprisingly, fragmented embryos, repaired or not, do implant and often come to term. This demonstrates the highly robust [regulative] nature of the human embryo, as it can apparently lose over half of its cellular mass and still recover." (p. 197) [Back]

3 Bruce M. Carlson, Human Embryology and Developmental Biology (St. Louis, MO: Mosby, 1999): "Early mammalian embryogenesis is considered to be a highly regulative process. Regulation is the ability of an embryo or an organ primordium to produce a normal structure if parts have been removed or added. At the cellular level, it means that the fates of cells in a regulative system are not irretrievably fixed and that the cells can still respond to environmental cues." (p. 44). "... Blastomere removal and addition experiments have convincingly demonstrated the regulative nature (i.e., the strong tendency for the system to be restored to wholeness) of early mammalian embryos. Such knowledge is important in understanding the reason exposure of early human embryos to unfavorable environmental influences typically results in either death or a normal embryo." (p. 46) [Back]

4  Carlson 1999: "Some types of twinning represent a natural experiment that demonstrates the highly regulative nature of early human embryos, ... ." (p. 48) "... Monozygotic twins and some triplets, on the other hand, are the product of one fertilized egg. They arise by the subdivision and splitting of a single embryo. Although monozygotic twins could ... arise by the splitting of a two-cell embryo, it is commonly accepted that most arise by the subdivision of the inner cell mass in a blastocyst. Because the majority of monozygotic twins are perfectly normal, the early human embryo can obviously be subdivided and each component regulated to form a normal embryo." (p. 49)

William Larsen, Essentials of Human Embryology (New York: Churchill Livingstone, 1998): "If the splitting occurred during cleavage -- for example, if the two blastomeres produced by the first cleavage division become separated -- the monozygotic twin blastomeres will implant separately, like dizygotic twin blastomeres, and will not share fetal membranes. Alternatively, if the twins are formed by splitting of the inner cell mass within the blastocyst, they will occupy the same chorion but will be enclosed by separate amnions and will use separate placentae, each placenta developing around the connecting stalk of its respective embryo. Finally, if the twins are formed by splitting of a bilaminar germ disc, they will occupy the same amnion." (p. 325)

O'Rahilly and Muller 2001: "The embryo enters the uterine cavity after about half a week ... Each cell (blastomere) is considered to be still totipotent (capable, on isolation, of forming a complete embryo), and separation of these early cells is believed to account for one-third of cases of monozygotic twinning." (p.37) [Back]

5  Many IVF providers are strongly promoting the use of twinning cloning techniques as forms of "fertility treatments", a process they refer to as "embryo multiplication", e.g.: "Because early embryonic cells are totipotent, the possibility of splitting or separating the blastomeres of early preimplantation embryos to increase the number of embryos that are available for IVF treatment of infertility is being discussed. Because embryo splitting could lead to two or more embryos with the same genome, the term "cloning" has been used to describe this practice. ... Whereas these ethical concerns raise important issues, neither alone nor together do they offer sufficient reasons for not proceeding with research into embryo splitting and blastomere separation. ... In sum, since embryo splitting has the potential to improve the efficacy of IVF treatments for infertility, research to investigate the technique is ethically acceptable. Persons asked to donate gametes or embryos for such research should be fully informed that research in embryo splitting is intended or planned as a result of their donation. The fears of possible future abuses of the technique are not sufficient to stop valid research in use of embryo splitting as a treatment for infertility. This statement was developed by the American Society for Reproductive Medicine's Ethics Committee and accepted by the Board of Directors on December 8, 1995." See, AMERICAN SOCIETY OF REPRODUCTIVE MEDICINE.

See also: "New Ways to Produce Identical Twins -- A Continuing Controversy": "Now, a new method of actually producing identical twins looms near. Called "blastomere separation" (the separation of a two- to eight-cell blastomere into two identical demi-embryos), it is potentially one method of helping infertile couples have children through in vitro fertilization (IVF). ... The following is excerpted from the medical journal Assisted Reproduction Reviews, May 1994. Dr. Joe B. Massey, who heads an in vitro clinic in Atlanta. Dr. Massey reviews the advances in blastomere separation and discusses the potential indications, benefits, limitations, and ethics of using this method to produce monozygotic twin embryos for IVF patients. The Twins Foundation, by presenting Dr. Massey's material for your information neither advocates nor rejects any such procedures: 'Embryo Multiplication by Blastomere Separation -- One Doctor's Proposal'. [Massey]: In spite of many advances in human in vitro fertilization (IVF), there are still many problems. ... According to Dr. Massey, 'Observations on the potential impact of removing less than half of the cells from the human embryo have been well documented in pre-clinical embryo biopsy studies.' (For more on this story see Research Update Vol. 9, No. 1, 1994)." See, THE TWINS FOUNDATION.

See also: Professor Dr. Mithhat Erenus, "Embryo Multiplication": "In such cases, patients may benefit from embryo multiplication, as discussed in the study by Massey and co-workers. ... Since each early embryonic cell is totipotent (i.e., has the ability to develop and produce a normal adult), embryo multiplication is technically possible. ... In humans, removal of less than half of the cells from an embryo have been documented. No adverse effects were reported when an eighth to a quarter of the blastomeres were removed from an embryo on day 3 after insemination. ... Further evidence supporting the viability and growth of partial human embryos is provided by cryopreservation. After thawing four-cell embryos, some cells may not survive, leaving one-, two-, or three-cell embryos. These partial embryos survive and go to term, but at a lower rate than whole embryos. ... Based on the results observed in lower order mammals, the critical period of development to ensure success in separating human blastomeres should be at the time of embryonic gene expression, which is reported in humans to be between the four- and eight-cell stages [twinning by "blastomere splitting"]. .... The second potential method of embryo multiplication is blastocyst splitting. ... For couples who have less than three quality embryos for transfer, blastomere separation could be of benefit.." at: http://www.hekim.net/~erenus/20002001/asistedreproduction/micromanipulation/embryo_multiplication.htm.

Also, ETHICS COMMITTEE OF THE AMERICAN SOCIETY FOR REPRODUCTIVE MEDICINE, "'Ethical Considerations of Assisted Reproductive Technologies': Originally published as a supplement to the ASRM medical journal (Fertility and Sterility 1994;62:Suppl 1), Ethical Considerations for Assisted Reproductive Technologies covers the American Society for Reproductive Medicine's position on several aspects of reproductive medicine, including: ... the moral and legal status of the preembryo, ... the use of donor sperm, donor oocytes and donor preembryos, ... the cryopreservation of oocytes and preembryos, micro techniques such as: zona drilling, microinjection, blastomere separation (cloning), and assisted hatching." at: http://www.asrm.com/Media/Ethics/ethics94.html. [Back]

6 For over 30 pages of current scientific references documenting this, see Irving, "Framing the Debates on Human Cloning and Human Embryonic Stem Cells: Pluripotent vs. TOTIPOTENT" (July 23, 2005), at: http://www.lifeissues.net/writers/irv/irv_100debatecloning1.html. [Back]

7 Gao S, Sun W, Wen J, Guan Q, Zhang Q. (Reproductive Medicine Center, Jinan Central Railway Hospital, Jinan 250001, China), "Investigation on development potential of blastomeres isolated from 4-cell human embryos", Zhonghua Fu Chan Ke Za Zhi. 2002 Mar;37(3):155-6, PMID = 11953084, at: http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=pubmed&dopt=Abstract&list_uids=11953084&query_hl=5. [Back]

8  See, e.g., Meng QG, Zhu SE, Zeng SM, Zhang ZC, Li SR, Wang DP, Li JQ. (College of Animal Science, China Agricultural University, Beijing 100094), "Study on in vitro culture and cryopreservation by vitrification of blastocysts derived from single mouse 2-cell embryos' blastomeres", Shi Yan Sheng Wu Xue Bao. 2001 Dec;34(4):329-31, PMID: 12549214, at: http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=pubmed&dopt=Abstract&list_uids=12549214&query_hl=5.

Also, Taniguchi T, Taniguchi R, Kanagawa H. (Department of Theriogenology, Faculty of Veterinary Medicine, Hokkaido University, Sapporo, Japan), "Influence of oocyte aging on developmental ability of reconstituted embryos produced from oocyte cytoplast and single blastomeres of two-cell stage embryos", J Vet Med Sci. 1996 Jul;58(7):635-40, PMID: 8844599, at: http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=pubmed&dopt=Abstract&list_uids=8844599&query_hl=2. [Back]

9 Tarkowski AK, Ozdzenski W, Czolowska R. (Department of Embryology, Institute of Zoology, Warsaw University, Poland. AKT@biol.uw.edu.pl), "Mouse singletons and twins developed from isolated diploid blastomeres supported with tetraploid blastomeres", IntJ Dev Biol. 2001;45(3):591-6, PMID: 11417903, [full text available; research study dedicated to Ann McClaren, British biologist who coined the term "pre-embryo" for the Warnock Commission 1985], at: http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=pubmed&dopt=Abstract&list_uids=11417903&query_hl=5 [Back]